Fig. 5

The enzymatic activity of PYGM is required for muscle cell differentiation. A Representative images of immunostaining for MyoG (green) or MHC (green) in C2C12 cells treated with 100 μM of PYGM activity inhibitor (C5H11NO3·HCl, Sigma, D1542) and cultured in differentiation medium for 24 h (MyoG) or 48 h (MHC). DAPI (pseudo-colored red) served to visualize nuclei. Scale bars, 100 μm. B Numbers of MyoG-positive (MyoG⁺) cells per view described in (A). C Relative expression of MyoG in the cells described in (A), as determined by RT–qPCR. D Fusion index calculated in the cells described in (A). E Representative images of immunostaining for MyoG (green) or MHC (green) on C2C12 cells transfected with plasmids expressing wild-type (WT) or mutant PYGM (S14A) and induced to differentiate for 24 h (MyoG) or 48 h (MHC). Transfection with empty vector served as the negative control (NC). DAPI (pseudo-colored red) served to visualize nuclei. Scale bars, 100 μm. F Numbers of MyoG-positive (MyoG⁺) cells per view described in (E). G Relative expression of MyoG in the cells described in (E), as determined by RT–qPCR. H Fusion index calculated in the cells described in (E). All images are representatives of three independent experiments. Values presented represent the means ± SE obtained from three independent experiments. The statistical significance of the difference between two means was calculated with the Student’s t-test. *p < 0.05, **p < 0.01, ***p < 0.001